Gastrointestinal cancers of the microsatellite mutator phenotype (MMP) differ in genotype and phenotype from tumors without the MMP. APC, p53 and K-ras cancer genes are commonly mutated in MMP- tumors while mutational inactivation of TGFbetaRII and BAX is essentially restricted to MMP+ cancers. We propose that the existence and distinctive features of the MMP pathway for cancer, characteristic of tumors of the Hereditary Non-Polyposis Colorectal Cancer (HNPCC) syndrome and about 15% of sporadic gastrointestinal cancers, ultimately depends on the presence in some cancer genes of sequences target for the MMP. In specific aim 1 we will test the hypothesis of the existence of a MMP-dependent pathway for cancer. We propose to investigate the differences in genotype between tumors with or without the MMP. We will complete the screening of mutations in the APC suppressor gene in MMP+ and MMP- colon tumors (1a); determine whether the differences in genotype between MMP+ and MMP- gastrointestinal tumors also extend to widespread somatic changes in DNA methylation that we have observed in colon, breast and prostate cancer, specifically in the homeobox gene family (1b); and exploit the peculiar phenotypic features of MMP cancer to develop diagnostic assays for hereditary and sporadic cancer of the MMP (1c). In specific aim 2 we will test the hypothesis that the MMP unfolds gradually by the mutational inactivation of multiple mutator genes. We propose to investigate the model of the "mutator that mutates another mutator". We will complete the screening of colorectal and gastric MMP+ adenocarcinomas for mutations in the known members of the DNA mismatch repair (MMR) gene family (2a); perform a functional analysis of the effect of mutations in individual MMR mutator genes and their combinations in the spectra of mutations in the tumor cells (2b); and establish the prognostic value of these findings for cancer of the MMP (2c). In specific aim 3 we will test the hypothesis that the escape from apoptosis is a critical event in tumorigenesis of the MMP. We propose to investigate the mechanisms by which BAX mutational inactivation contributes to the escape from apoptosis and to tumorigenesis of tumors cells of the MMP. We will complete the screening of MMP+ and MMP- tumors for mutations in BAX (3a); perform a functional analysis of the role of BAX gene inactivation in cancer of the MMP by in vivo and in vitro assays (3b); and investigate the prognostic value of BAX somatic mutational inactivation for cancer of the MMP.